JP2021036228A - Micro biosensor and measuring method thereof - Google Patents

Abstract

To provide a measuring method for prolonging a usage lifetime of a micro biosensor to measure a physiological signal associated with an analyte.SOLUTION: A micro biosensor includes a working electrode, a counter electrode including silver and a silver halide having an initial amount, and an auxiliary electrode. The method includes cyclic steps of: applying a measurement voltage to drive the working electrode and measure a physiological signal to obtain a physiological parameter, where the silver halide is consumed by a specific amount; stopping applying the measurement voltage; and whenever the physiological parameter is obtained, applying a replenishment voltage between the counter electrode and the auxiliary electrode to drive the counter electrode, thereby an oxidation reaction being caused and the silver halide of a replenishment amount being replenished to the counter electrode. A guarding value of a sum of the replenishment amount and the initial amount subtracting the consumption amount is controlled within a range of the initial amount plus or minus a specific value.SELECTED DRAWING: Figure 10

Description

Cross-reference between related application and priority claim <br /> This application is a US provisional patent application filed on August 2, 2019, US provisional patent application number 62 / 882,162 and a US provisional patent application filed on March 12, 2020. Claiming the interests of filing date No. 62 / 988,549, these disclosures are incorporated herein by reference in their entirety.

The present invention relates to a microbiosensor and a measuring method thereof, and more particularly to a microbiosensor for extending the service life of the microbiosensor and a measuring method thereof.

The population of diabetics is growing rapidly, and the need to monitor changes in glucose in the human body is increasingly emphasized. Therefore, in order to eliminate the inconvenience of patients due to repeated blood sampling and detection every day, many studies have begun to develop a continuous blood glucose monitoring (CGM) system that can be implanted in the human body.

In the field of enzyme-based biosensors of CGM systems, biochemical reaction signals that depend on the concentration of the analyte are converted into measurable physical signals such as optical or electrochemical signals. In the case of glucose measurement, an electrochemical reaction occurs and glucose oxidase (GOx) catalyzes glucose to react to produce gluconolactone and reductase. Next, the reductase transfers electrons to oxygen in the biofluid in the living body to generate hydrogen peroxide (H ₂ O ₂ ), which is a product, and oxidizes the product H 2 O 2 to quantify the concentration of glucose. To become.
The reaction is as follows.
Glucose + GOx (FAD) → GOx (FADH2) + Gluconolactone GOx (FADH2) + O ₂ → GOx (FAD) + H ₂ O ₂
Here, FAD (flavin adenine dinucleotide) is the active center of GOx.

Since users usually wear a CGM system for a long period of time (for example, at least 14 days), reducing its size is a necessary development. The basic structure of the CGM system includes (a) a biosensor that measures physiological signals corresponding to the glucose concentration of the human body, and (b) a transmitter for transmitting these physiological signals. The biosensor can be a two-electrode system or a three-electrode system. A biosensor equipped with a three-electrode system includes a working electrode (WE), a counter electrode (CE) and a reference electrode (RE). A biosensor with a two-electrode system includes a working electrode (WE) and a counter electrode (CE), which also functions as a reference electrode and is therefore also referred to as a reference electrode / counter electrode (R / C). is there. For the reference electrode of the biosensor of the three-electrode system and the counter electrode that also functions as the reference electrode of the biosensor of the two-electrode system, the appropriate materials applicable for stable measurement of glucose concentration are silver and silver chloride (Ag / AgCl). ). However, after the biosensor is implanted in the living body, an oxidation reaction occurs at the working electrode, and when the glucose concentration is measured, a reduction reaction occurs at the corresponding reference electrode (R) or reference electrode / counter electrode (R / C). Causes, reduces AgCl to Ag, and consumes AgCl. Furthermore, when the biosensor embedded in the living body is a biosensor of a two-electrode or three-electrode system, silver chloride dissociates in the body fluid, which consumes silver chloride from the reference electrode and causes a drift problem with respect to the reference electrode. However, due to the reaction of the reference electrode / counter electrode (R / C) of the two-electrode system, the consumption of silver chloride is even higher than that of the three-electrode system. Therefore, the useful life of the biosensor is limited by the silver chloride content on the counter electrode and / or the reference electrode.

There are also many inventions proposed to address this issue. Taking a biosensor equipped with a two-electrode system as an example, the consumption at the counter electrode is about 1.73 millicoulombs (mC) per day with an average detection current of 20 nanoamperes (nA). Assuming that the length, width and height of the counter electrode are 3.3 mm, 0.25 mm and 0.01 mm, respectively, and the initially designed electrode capacitance is 6 mC, the maximum stability measurement that the biosensor can provide is about 1. Can be maintained for a day. However, the counterelectrode volume should be at least 27.68 mC if the biosensor's useful life needs to be further extended so that the subcutaneously implanted biosensor can support 16 days of continuous blood glucose monitoring. .. Without changing the width and thickness of the counter electrode, the length of the counter electrode of the prior art should be up to 15.2 mm. Therefore, the length of the counter electrode of the biosensor is extended to be greater than 10 mm in the prior art. However, to avoid deep implantation of such types of biosensors in the subcutaneous tissue, the biosensors need to be implanted at an oblique angle. Therefore, problems such as an increase in the size of the implanted wound and an increased risk of infecting the patient occur, and since the implantation length is long, the pain during implantation also increases.

U.S. Pat. No. 8,620,398 primarily describes biosensors with a three-electrode system. The reference electrode is basically not involved in the chemical reaction, but silver chloride is naturally and gradually consumed in the in vivo environment, and the consumption rate is slower than that of the counter electrode of the two-electrode system. The specification discloses that AgCl regenerates when it is almost completely depleted. That is, the replenishment process is run until the measurement signal becomes unstable, or until the measurement signal is all noisy, and AgCl recovers to a sufficient amount to perform multiple measurements. Then, AgCl needs to be replenished again until the next noise is generated. U.S. Pat. No. 8,620,398 believes that if a biosensor fails, AgCl will be depleted during measurement and replenishment of AgCl, but it is understandable that the measurements at failure are no longer reliable. In order to obtain the correct reading, it is necessary to wait for the biosensor to complete the AgCl replacement process, take a blood sample and perform the measurement temporarily, or skip this measurement directly. This problem is always troublesome for patients and those who need to know their current blood glucose levels. In addition, the biosensor needs to process multiple measurements in a row or multiple measurements over several days, so more AgCl volumes need to be prepared. However, it will inevitably lead to the problem of longer implantation lengths of biosensors. U.S. Pat. No. 8,620,398 does not propose any timely AgCl replenishment method that can provide uninterrupted measurements and shorter implantation lengths and longer service lives of biosensors.

US9,351,677 proposes a sensor for measuring the analyte, which mainly uses a two-electrode system. Since the reference electrode / counter electrode (R / C) is involved in the chemical reaction, silver chloride is consumed by the electrochemical reaction. This patent discloses an analyte sensor with increased AgCl volume. The sensor uses H2O2 to regenerate AgCl on the reference electrode. However, since H2O2 is easily reduced to H2O or oxidized to O2, it is not easy to stably exist in the human body. Therefore, during the regeneration / replenishment period, the H2O2 concentration in the human body may not be sufficient to stably replenish a sufficient amount of AgCl, and the biosensor needs to be equipped with a larger AgCl electrode size. Yes, the embedded end has a maximum length of 12 mm.

Therefore, the present disclosure provides uninterrupted measurement by replenishing AgCl after measurement, stably replenishes AgCl, prolongs the life of the biosensor, and compacts the embedded end of the biosensor. It provides a biosensor that can achieve the effect of reducing the size and manufacturing cost of the product. These effects can solve the above-mentioned problems that the prior art has found insurmountable.

In view of the above, due to the deficiencies of the prior art, we provide the present invention to effectively overcome the disadvantages of the prior art. The description of the present invention is as follows.

According to the replenishment technique of the present invention, the microbiosensor of the present invention has a long service life, the size of the signal sensing portion of the counter electrode of the microbiosensor can be reduced, and the biotoxicity can be reduced. In addition, reducing the size of the electrodes specifically refers to the shortened length of the embedded end of the sensor, which will reduce user pain during implantation.

According to another aspect of the present disclosure, a biosensor implanted subcutaneously to measure a physiological signal representing a physiological parameter associated with an analyte in a biofluid, for extending the useful life of the biosensor. Discloses a method of measuring the analyte using the biosensor. The biosensor includes a working electrode, a counter electrode and an auxiliary electrode, the working electrode is at least partially covered with a chemical reagent to react with the analyte, and the counter electrode is silver and silver halide. Have. The method is (a) a measurement step of carrying out the measurement, i, causing a first oxidation reaction at the working electrode having an electrochemical reaction with the chemical reagent and the analysis product to obtain the current physiological signal. In order to output, a step of applying a measurement potential difference between the working electrode and the counter electrode so that the working electrode has a voltage level higher than the voltage level of the counter electrode during the measurement period. The silver halide of the counter electrode has a sub-step having a current consumption amount corresponding to the current physiological signal, and ii, the measurement potential difference is removed to stop the measurement step, and the current physiological signal is calculated. In the measurement step including the sub-step for outputting the current physiological parameters, and (b) the replenishment step for executing the replenishment, i, the voltage at which the counter electrode is higher than the voltage level of the auxiliary electrode. A replenishment potential difference is applied between the counter electrode and the auxiliary electrode during the replenishment period to cause a carbon dioxide reaction on the silver on the counter electrode so that the halide silver is depleted. Obtaining a replenishment amount corresponding to the amount, the silver halide of the counter electrode has an amount maintained in a safe storage range, and the next physiological signal and the next physiological parameter obtained in the next measurement step are specified. The replenishment step including the sub-step that keeps the correlation of ii, the sub-step that removes the replenishment potential difference and stops the replenishment step, and (c) the next including the same sub-step as step (a). Includes a step of executing the measurement step of (d) and a step of executing the next replenishment step including the same substep as step (b).

According to another aspect of the present disclosure, a biosensor implanted subcutaneously to measure a physiological signal representing a physiological parameter associated with an analyte in a biofluid, extending the useful life of the biosensor. Therefore, a method of measuring the analysis product using the biosensor is disclosed. It comprises the biosensor, working electrode, counter electrode and auxiliary electrode, the working electrode is at least partially covered with a chemical reagent, and the counter electrode contains silver and silver halide and has an initial amount. In the method, a measuring voltage is applied to drive the working electrode, and the physiological signal is measured to acquire the physiological parameters, and the silver halide is consumed by the consumption amount, and the measured voltage is measured. The step of stopping the application and each time the physiological parameter is acquired, a replenishment voltage is applied between the counter electrode and the auxiliary electrode to drive the counter electrode and cause an oxidation reaction to cause the replenishment amount. The guard value obtained by subtracting the consumable amount from the sum of the replenishment amount and the initial amount, including the step of replenishing the counter electrode with silver halide, is controlled within a range in which a specific value is increased or decreased from the initial amount. Will be done.

According to another aspect of the present disclosure, a microbiosensor for subcutaneous implantation is provided for measuring physiological parameters representing physiological signals associated with an in vivo analyte. The microbiosensor is placed on the substrate, the chemical reagent, and the substrate, at least partially covered with the chemical reagent, and driven for the first oxidation reaction within the measurement period to measure the physiological signal. A working electrode for acquiring the physiological parameters and a counter electrode arranged on the substrate and containing silver and silver halide, wherein the silver halide has an initial amount and is consumed within the measurement period. The counter electrode that is consumed by the amount and the auxiliary electrode that is arranged on the substrate, and each time the physiological parameters are acquired, the counter electrode and the auxiliary electrode are subjected to the carbon dioxide reaction within the replenishment period. The guard value obtained by subtracting the consumable amount from the sum of the replenishment amount and the initial amount is the guard value including the auxiliary electrode in which the replenishment amount of the silver halide is replenished to the counter electrode. It is controlled within the range in which a specific value is increased or decreased from the initial amount.

The above embodiments and advantages of the present invention will become more apparent to those skilled in the art after reviewing the following detailed description and accompanying drawings.
The schematic diagram of the physiological signal measuring apparatus of this invention is shown. The front schematic view of the 1st Embodiment of the microbiosensor of this invention is shown. The back schematic of the 1st Embodiment of the microbiosensor of this invention is shown. A schematic cross-sectional view of the microbiosensor along the cross-sectional lines A to A'of FIG. 2A is shown. A schematic cross-sectional view of the second embodiment of the microbiosensor of the present invention is shown. A schematic cross-sectional view of a third embodiment of the microbiosensor of the present invention is shown. A schematic cross-sectional view of a fourth embodiment of the microbiosensor of the present invention is shown. A schematic cross-sectional view of a fifth embodiment of the microbiosensor of the present invention is shown. A schematic cross-sectional view of the sixth embodiment of the microbiosensor of the present invention is shown. A schematic cross-sectional view of a seventh embodiment of the microbiosensor of the present invention is shown. A schematic cross-sectional view of the eighth embodiment of the microbiosensor of the present invention is shown. The constant voltage circuit in the measurement mode in this invention is shown. The constant voltage circuit of the replenishment mode in this invention is shown. The schematic diagram of the current of the constant voltage circuit which operates in the measurement mode and the replenishment mode alternately by the 1st method is shown. The schematic diagram of the current of the constant voltage circuit which operates in the measurement mode and the replenishment mode alternately by the 2nd method is shown. The schematic diagram of the current of the constant voltage circuit which operates in the measurement mode and the replenishment mode alternately by the 3rd method is shown. The schematic diagram of the current of the constant voltage circuit which operates in the measurement mode and the replenishment mode alternately by the 4th method is shown. The schematic diagram of the current of the constant voltage circuit which operates in the measurement mode and the replenishment mode alternately by the 5th method is shown. The schematic diagram of the current of the constant voltage circuit which operates in the measurement mode and the replenishment mode alternately by the 6th method is shown. The segment constant current circuit in the measurement mode in this invention is shown. The segment constant current circuit of the replenishment mode in this invention is shown. The continuously variable constant current circuit in the measurement mode in this invention is shown. The continuously variable constant current circuit of the replenishment mode in this invention is shown. The voltage schematic diagram of the constant current circuit which operates in the measurement mode and the replenishment mode alternately by the 1st method is shown. The voltage schematic diagram of the constant current circuit which operates in the measurement mode and the replenishment mode alternately by the 2nd method is shown. The voltage schematic diagram of the constant current circuit which operates in the measurement mode and the replenishment mode alternately by the 3rd method is shown. The schematic diagram of the constant current circuit which operates in the measurement mode and the replenishment mode alternately by the 3rd method is shown. A method for measuring an analytical object according to an embodiment of the present invention is shown. A method for measuring an analyzer according to another embodiment of the present invention is shown.

All figures of the present invention are referred to when reading the detailed description below. All figures of the invention show different embodiments of the invention by illustration to help one of ordinary skill in the art to understand how to practice the invention. The present embodiment provides sufficient embodiments to demonstrate the spirit of the present invention, each embodiment is consistent with other embodiments, and new embodiments may be implemented through any combination thereof. it can. That is, the present invention is not limited to the embodiments disclosed herein.

Unless otherwise restricted as defined in a particular example, the following definitions apply to terms used throughout the specification.

The term "amount" refers to the volume of silver halide (AgX) or silver chloride (AgCl) on the counter electrode, preferably in units of microcoulomb (μC), millicoulomb (mC) or coulomb (C). Represent. However, the concentration is not limited to the weight percentage (wt%), the number of moles, the molar concentration, and the like.

With reference to FIG. 1, FIG. 1 is a schematic view of the physiological signal measuring device of the present invention. The physiological signal measuring device 10 of the present invention can be implanted subcutaneously to measure a physiological signal representing a physiological parameter associated with an analyte in a biological fluid.
The physiological signal measuring device 10 of the present invention includes a microbiosensor 300 and a transmitter 200, and the transmitter 200 is electrically connected to the microbiosensor 300 and has a processor 210, a power supply 220, a circuit switching unit 230, and a temperature sensing device. Includes unit 240 and communication unit 250. The power supply 220 provides a voltage to the microbiosensor 300 via the circuit switching unit 230 to measure the physiological signal. The temperature sensing unit 240 measures the body temperature of the living body and transmits the temperature measurement signal and the physiological signal measured by the microbiosensor 300 to the processor 210. The processor 210 calculates the measured physiological signal with respect to the physiological parameter. The communication unit 250 can communicate with the user device 20 by wire or wireless transmission.

With reference to FIGS. 2A and 2B, FIGS. 2A and 2B are schematic front and back views of a first embodiment of the microbiosensor of the present invention, respectively. The microbiosensor 300 of the present invention includes a substrate 310, a working electrode 320, a counter electrode 330 arranged on the substrate 310, and a chemical reagent 350 covering the working electrode 320, the counter electrode 330, and the auxiliary electrode 340 (see FIG. 2C). Show) and includes. The material of the substrate 310 can be any material that is suitable for use in electrode substrates and is known to have flexibility and insulating properties, preferably polymer materials such as polyester and polyimide. Not limited. The above-mentioned polymer materials can be used alone or in combination. The substrate 310 includes a surface 311 (ie, the first surface), a surface 312 opposite the surface 311 (ie, the second surface), a first end 313, and a second end 314. The substrate 310 is arranged between the signal output region 315 arranged near the first end portion 313, the sensing region 316 arranged near the second end portion 314, and the signal output region 315 and the sensing region 316, respectively. It is separated into three areas with the connection area 317 to be formed. The working electrode 320 is arranged on the surface 311 of the substrate 310 and extends from the first end 313 to the second end 314 of the substrate 310. The working electrode 320 includes a signal output portion 321 arranged in the signal output region 315 of the substrate 310 and a signal sensing portion 322 arranged in the sensing region 316 of the substrate 310.

The counter electrode 330 and the auxiliary electrode 340 are arranged on the surface 312 on the opposite side of the substrate 310 and extend from the first end 313 to the second end 314 of the substrate 310. The counter electrode 330 includes a signal sensing portion 332 arranged in the sensing region 316 of the substrate 310, and the auxiliary electrode 340 includes a signal sensing portion 342 arranged in the sensing region 316 of the substrate 310. The sensing region 316 of the microbiosensor 300 can be implanted subcutaneously so that the signal sensing portion 322 can measure the physiological signal of the analyte in the biological fluid. The physiological signal is transmitted to the processor 210 via the signal output portion 321 in order to obtain the physiological parameters. Further, apart from the transmitter 200, physiological parameters can also be acquired from the user device 20 via wired / wireless communication. The general user device 20 can be a smartphone, a physiological signal receiver, or a glucose meter.

The surface material of the counter electrode 330 contains silver and silver halide, preferably silver chloride or silver iodide. Since the electrode material of the counter electrode 330 of the present invention contains silver and silver halide (Ag / AgX), the counter electrode 330 of the present invention includes the functions of the counter electrode and the reference electrode well known in the art. Specifically, the counter electrode 330 of the present invention (1) forms an electronic circuit with the working electrode 320, conducts a current between the counter electrode 330 and the working electrode 320, and causes an oxidation reaction at the working electrode 320. Can be ensured that (2) provides a stable relative potential as a reference potential. Therefore, the working electrode 320 and the counter electrode 330 of the present invention form a two-electrode system. Ag / AgX can be used with carbon to further reduce the cost of the biosensor of the invention and improve biocompatibility, eg, Ag / AgX is mixed with a carbon paste and contains silver halide. The amount is an amount at which the counter electrode 330 can stably perform the measurement step. The surface of the counter electrode 330 may be partially covered with a conductive material to prevent the silver halide from dissociating and to protect the counter electrode 330. Here, the conductive material is selected from materials that do not affect the measurement result of the working electrode, for example, the conductive material is carbon.

In another embodiment, the biosensor is not limited to a wire type or stack type electrode structure.

According to another embodiment of the disclosure, the initial amount of silver halide can be zero before the biosensor is ready to be shipped from the factory for sale. In this case, the counter electrode 330 of the biosensor does not have silver halide. The initial amount of silver halide can be replenished by oxidizing the silver coated on the counter electrode 330 after the biosensor has been subcutaneously implanted in the patient and during the first replenishment period prior to the first measurement. ..

The auxiliary electrode 340 forms an electronic circuit with the counter electrode 330 in the replenishment step and conducts an electric current between the counter electrode 330 and the auxiliary electrode 340 to ensure that an oxidation reaction occurs on the counter electrode 330. The electrode material of the auxiliary electrode 340 may be the same as the electrode material of the working electrode 320, or the sensitivity to hydrogen peroxide such as carbon may be lower than that of the working electrode 320.

The chemical reagent 350 covers at least a partial surface of the signal sensing portions 322, 332, 342 of each electrode. In another embodiment, the chemical reagent 350 covers at least a partial surface of the signal sensing portion 322 of the working electrode 320 (not shown). Specifically, the counter electrode 330 is not covered by the chemical reagent 350. The sensing region 316 of the microbiosensor 300 is implanted subcutaneously so that the signal sensing portion 322 of the working electrode 320 can measure the physiological signal of the analyte in the biological fluid. The physiological signal is transmitted to the processor 210 via the signal output portion 321 of the working electrode 320 to acquire the physiological parameters. Further, apart from the transmitter 200, physiological parameters can also be acquired from the user device 20 via wired / wireless communication.

With reference to FIG. 2C, FIG. 2C is a schematic cross-sectional view of the microbiosensor along the cross-sectional lines A to A'of FIG. 2A. In FIG. 2C, lines A to A'are cross-sectional lines of the sensing region 316 of the microbiosensor 300.
In FIG. 2C, the working electrode 320 is placed on the surface 311 of the substrate 310, the counter electrode 330 and the auxiliary electrode 340 are placed on the surface 312 opposite the substrate 310, the working electrode 320, the counter electrode 330 and the auxiliary. The surface of the electrode 340 is covered with the chemical reagent 350. Basically, the chemical reagent 350 covers at least a partial surface of the working electrode 320. The microbiosensor 300 of the present invention performs the measurement step during the measurement period and performs the replenishment step during the replenishment period. When the measurement step is performed, the voltage level of the working electrode 320 becomes higher than the voltage level of the counter electrode 330, and a current flows from the working electrode 320 to the counter electrode 330, resulting in an electrochemical reaction with the chemical reagent 350 and the analyte. An oxidation reaction occurs on the working electrode 320, the physiological signal is measured, and a reduction reaction occurs on the counter electrode 330, and as a result, the silver halide (AgX) in the counter electrode 330 is consumed and becomes silver (Ag). It is dissociated into a halogen ion (X-). Since the silver halide in the counter electrode 330 is consumed, it is necessary to replenish the silver halide in the counter electrode 330 in order to carry out the next measurement step. When the replenishment step is performed, the voltage level of the counter electrode 330 becomes higher than the voltage level of the auxiliary electrode 340, and a current flows from the counter electrode 330 to the auxiliary electrode 340, resulting in an oxidation reaction at the counter electrode 330 and silver. It replenishes silver halide by binding with halogen ions in the living body. A detailed measurement step and a detailed replenishment step are shown in FIG.

With reference to FIG. 3A, FIG. 3A is a schematic cross-sectional view of a second embodiment of the microbiosensor of the present invention. In FIG. 3A, the working electrode 320 and the auxiliary electrode 340 may be arranged on the surface 311 of the substrate 310, the counter electrode 330 is arranged on the surface 312 opposite the substrate 310, and the working electrode 320, the counter electrode. The surfaces of 330 and the auxiliary electrode 340 are covered with the chemical reagent 350. In this embodiment, when the measurement step is performed, a current flows from the working electrode 320 to the counter electrode 330, resulting in an oxidation reaction occurring on the working electrode 320, the physiological signal being measured and the silver halide at the counter electrode 330. AgX) is consumed, silver (Ag) and silver ions (X ^- is dissociated). When the replenishment step is executed, a current flows from the counter electrode 330 to the auxiliary electrode 340, an oxidation reaction occurs on the counter electrode 330, and silver halide is replenished by a combination of silver ions and halogen ions.

With reference to FIG. 3B, FIG. 3B is a schematic cross-sectional view of a third embodiment of the microbiosensor of the present invention. In this embodiment, the microbiosensor 300 has two working electrodes, a first working electrode 323 and a second working electrode 324, respectively. Here, the auxiliary electrode is replaced by a second working electrode 324. In FIG. 3B, the first working electrode 323 and the second working electrode 324 are arranged on the surface 311 of the substrate 310, and the counter electrode 330 is arranged on the surface 312 opposite the substrate 310, the first working electrode. The surfaces of 323, the second working electrode 324 and the counter electrode 330 are covered with the chemical reagent 350. One of the first working electrode 323 and the second working electrode 324 was selected to measure the physiological signal in the measurement step, and the first working electrode 323 or the second working electrode 324 was silver halide in the replenishment step. To replenish the counter electrode 330, an electronic circuit is formed with the counter electrode 330. Therefore, in this embodiment, when the measurement step is performed, a current flows from the first working electrode 323 or the second working electrode 324 to the counter electrode 330. As a result, an oxidation reaction occurs at the first working electrode 323 or the second working electrode 324, the physiological signal is measured, the silver halide in the counter electrode 330 (AgX) is consumed, silver (Ag) and silver ions (X ^- ) Is dissociated. When the replenishment step is performed, a current flows from the counter electrode 330 to the first working electrode 323 or the second working electrode 324. As a result, an oxidation reaction occurs on the counter electrode 330 to replenish silver halide with a combination of silver and halogen ions.

With reference to FIG. 3C, FIG. 3C is a schematic cross-sectional view of a fourth embodiment of the microbiosensor of the present invention. In this embodiment, the microbiosensor 300 has two working electrodes, a first working electrode 323 and a second working electrode 324, respectively. Here, the auxiliary electrode is replaced by a second working electrode 324. In FIG. 3C, the first working electrode 323 is placed on the surface 311 of the substrate 310, the counter electrode 330 and the second working electrode 324 are placed on the surface 312 opposite the substrate 310, and the first working electrode 323. The surfaces of the second working electrode 324 and the counter electrode 330 are covered with the chemical reagent 350. In this embodiment, the surface area of the first working electrode 323 can be increased to perform the measurement step and the second working electrode to perform the replenishment step to replenish the counter electrode 330 with silver halide. The surface area of 324 can be reduced. Therefore, in this embodiment, when the measurement step is performed, a current flows from the first working electrode 323 to the counter electrode 330. As a result, an oxidation reaction occurs at the first working electrode 323, the physiological signal is measured, the silver halide (AgX) is consumed in the counter electrode 330, silver (Ag) and silver ions ^- are dissociated into (X). When the replenishment step is performed, a current flows from the counter electrode 330 to the second working electrode 324. As a result, an oxidation reaction occurs on the counter electrode 330 to replenish silver halide with a combination of silver and halogen ions.

With reference to FIG. 3D, FIG. 3D is a schematic cross-sectional view of a fifth embodiment of the microbiosensor of the present invention. The fifth embodiment is the addition of another working electrode in the first embodiment. Specifically, the microbiosensor 300 has two working electrodes, respectively, a first working electrode 323 and a second working electrode 324, one counter electrode 330 and one auxiliary electrode 340. In FIG. 3D, the first working electrode 323 and the second working electrode 324 are arranged on the surface 311 of the substrate 310, and the counter electrode 330 and the auxiliary electrode 340 are arranged on the surface 312 on the opposite side of the substrate 310. The surfaces of the first working electrode 323, the second working electrode 324, the counter electrode 330 and the auxiliary electrode 340 are covered with the chemical reagent 350. In the measurement step, one of the first working electrode 323 and the second working electrode 324 is selected to measure the physiological signal, and in the replenishment step, the auxiliary electrode 340 forms an electronic circuit with the counter electrode 330. Then, silver halide is replenished to the counter electrode 330. Therefore, in this embodiment, when the measurement step is performed, a current flows from the first working electrode 323 or the second working electrode 324 to the counter electrode 330. As a result, an oxidation reaction occurs at the first working electrode 323 or the second working electrode 324, the physiological signal is measured, the silver halide (AgX) is consumed in the counter electrode 330, silver (Ag) and silver ions (X ^- ) Is dissociated. When the replenishment step is performed, a current flows from the counter electrode 330 to the auxiliary electrode 340. As a result, an oxidation reaction occurs on the counter electrode 330 to replenish silver halide with a combination of silver and halogen ions.

With reference to FIG. 3E, FIG. 3E is a schematic cross-sectional view of a sixth embodiment of the microbiosensor of the present invention. In this embodiment, the microbiosensor 300 has three working electrodes, the working electrodes 323, the second working electrode 324 and the third working electrode 325, respectively, which are auxiliary. The electrode is replaced by a third working electrode 325. In FIG. 3E, the first working electrode 323 and the second working electrode 324 are arranged on the surface 311 of the substrate 310, and the counter electrode 330 and the third working electrode 325 are arranged on the opposite surface 312 of the substrate 310. The surfaces of the first working electrode 323, the second working electrode 324, the third working electrode 325 and the counter electrode 330 are covered with the chemical reagent 350. One of the first working electrode 323, the second working electrode 324 and the third working electrode 325 is selected for measuring the physiological signal in the measuring step, the first working electrode 323, the second working electrode 324 or the first working electrode. The working electrode 325 forms an electronic circuit with the counter electrode 330 to replenish the counter electrode 330 with silver halide in the replenishment step. Therefore, in this embodiment, when the measurement step is performed, current flows from the first working electrode 323, the second working electrode 324 or the third working electrode 325 to the counter electrode 330. As a result, an oxidation reaction occurs at the first working electrode 323, the second working electrode 324 or the third working electrode 325, the physiological signal is measured, the silver halide (AgX) at the counter electrode 330 is consumed, and the silver (Ag) is consumed. Is dissociated into a halogen ion (X ^−). When the replenishment step is performed, current flows from the counter electrode 330 to the first working electrode 323, the second working electrode 324 or the third working electrode 325. As a result, an oxidation reaction occurs on the counter electrode 330 to replenish silver halide with a combination of silver and halogen ions.

With reference to FIG. 3F, FIG. 3F is a schematic cross-sectional view of a seventh embodiment of the microbiosensor of the present invention. The seventh embodiment is a modification of the electrode configuration of the sixth embodiment. In this embodiment, as shown in FIG. 3F, the first working electrode 323, the second working electrode 324 and the third working electrode 325 are arranged on the surface 311 of the substrate 310, and the counter electrode 330 is the opposite of the substrate 310. It is placed on the side surface 312. The surfaces of the first working electrode 323, the second working electrode 324, the third working electrode 325 and the counter electrode 330 are covered with the chemical reagent 350. One of the first working electrode 323, the second working electrode 324 and the third working electrode 325 is selected for measuring the physiological signal in the measuring step, the first working electrode 323, the second working electrode 324 or the first working electrode. The working electrode 325 forms an electronic circuit with the counter electrode 330 to replenish the counter electrode 330 with silver halide in the replenishment step. Therefore, in this embodiment, when the measurement step is performed, current flows from the first working electrode 323, the second working electrode 324 or the third working electrode 325 to the counter electrode 330. As a result, an oxidation reaction occurs at the first working electrode 323, the second working electrode 324 or the third working electrode 325, the physiological signal is measured, the silver halide (AgX) at the counter electrode 330 is consumed, and the silver (Ag) is consumed. Is dissociated into a halogen ion (X ^−). When the replenishment step is performed, current flows from the counter electrode 330 to the first working electrode 323, the second working electrode 324 or the third working electrode 325. As a result, an oxidation reaction occurs on the counter electrode 330 to replenish silver halide with a combination of silver and halogen ions.

With reference to FIG. 3G, FIG. 3G is a schematic cross-sectional view of an eighth embodiment of the microbiosensor of the present invention. Compared with FIG. 3D, the difference is that the second working electrode 324 of FIG. 3G is U-shaped. In the eighth embodiment, the first working electrode 323 and the second working electrode 324 are configured on the surface 311 of the substrate 310, and the second working electrode 324 is adjacent to and around the first working electrode 323. The counter electrode 330 and the auxiliary electrode 340 are arranged on the surface 312 on the opposite side of the substrate 310. Therefore, in this embodiment, when the measurement step is performed, current flows from the first working electrode 323 to the counter electrode 330. As a result, an oxidation reaction occurs at the first working electrode 323, the physiological signal is measured, the silver halide (AgX) is consumed in the counter electrode 330, silver (Ag) and silver ions ^- are dissociated into (X). When the replenishment step is performed, current flows from the counter electrode 330 to the auxiliary electrode 340 or the second working electrode 324. As a result, an oxidation reaction occurs on the counter electrode 330 to replenish silver halide with a combination of silver and halogen ions.

The detailed electrode stacks of FIGS. 2C-3G are omitted and only the electrode positions are shown.

In any of the above embodiments, the substrate 310 of the present invention is an insulator. The electrode materials of the working electrode 320 and the first working electrode 323 of the present invention include carbon, platinum, aluminum, gallium, gold, indium, iridium, iron, lead, magnesium, nickel, manganese, molybdenum, osmium, palladium, rhodium, and the like. Includes, but is not limited to, silver, tin, titanium, zinc, silicon, zirconium, mixtures thereof, or derivatives thereof (such as alloys, oxides or metal compounds). Preferably, the material of the working electrode 320 and the first working electrode 323 is a noble metal, a noble metal derivative, or a combination thereof. More preferably, the working electrode and the first working electrode 323 are made of a platinum-containing material. As the material of the second working electrode 324 and the third working electrode 325, the elements as exemplified for the above working electrode 320 and the first working electrode 323 or derivatives thereof can be used. In another embodiment, the electrode material of the second working electrode 324 and the third working electrode 325 may be a material that is less sensitive to hydrogen peroxide than the sensitivity of the first working electrode 323, such as carbon.

Since the electrode material of the counter electrode 330 of the present invention contains silver and silver halide (Ag / AgX), the counter electrode 330 of the present invention includes the functions of the counter electrode and the reference electrode well known in the art. Specifically, the counter electrode 330 of the present invention (1) forms an electronic circuit with the working electrode 320, conducts a current between the counter electrode 330 and the working electrode 320, and is electrochemical at the working electrode 320. The reaction can be ensured. (2) An electronic circuit is formed with the auxiliary electrode 340 to conduct a current between the counter electrode 330 and the auxiliary electrode 340, so that the electrochemical reaction is carried out on the counter electrode 330. Provide a stable relative potential as a reference potential (3) to ensure that it occurs above. Therefore, the working electrode 320, the counter electrode 330, and the auxiliary electrode 340 of the present invention form a three-electrode system different from the conventional three-electrode system.

When the electrode material of the auxiliary electrode 340 of the present invention is covered with platinum, the auxiliary electrode 340 can also be used as an electrode for measuring a physiological signal.

In any of the above embodiments, a layer of conductive material, such as carbon, is placed between the surface 312 on the opposite side of the substrate 310 and the silver of the counter electrode 330 to prevent the silver electrode material from being damaged by excessive chlorination. Can be further placed. However, if the bottom layer of the counter electrode 330 is carbon, the resistance at the switch position is too high. A conductive layer such as silver can be further placed between the carbon conductive material and the surface 312 on the opposite side of the substrate 310. Therefore, the material of the counter electrode 330 of the present invention is a conductive layer, a carbon layer, and a silver / silver halide layer in this order from the surface 312 on the opposite side of the substrate 310.

Switching application of constant voltage circuit With reference to FIGS. 4A to 4B and 5A to 5D, FIGS. 4A and 4B show the constant voltage circuit in the measurement mode and the replenishment mode in the present invention, respectively. ~ D show in order a schematic diagram of the current of the constant voltage circuit which operates in the measurement mode and the replenishment mode alternately by different methods, respectively. The measurement mode can be started and stopped by applying the measurement potential difference V1 and removing the measurement potential difference V1, and the corresponding current is represented by Ia. In the measurement mode, since the measurement potential difference V1 is applied between the working electrode W and the counter electrode R / C during the measurement period T1, the voltage of the working electrode W becomes higher than the voltage of the counter electrode R / C. In the measurement mode, as shown in FIG. 4A, the switches S1 and S4 are in the closed circuit state, the switches S2 and S3 are in the open circuit state, the working electrode W is + V1, and the auxiliary electrode Aux is in the open circuit state. The counter electrode R / C is grounded. As a result, an oxidation reaction occurs at the working electrode W, and the working electrode W electrochemically reacts with the chemical reagent and the analyte to output the physiological signal Ia. AgCl in the counter electrode R / C has a consumable amount corresponding to the physiological signal Ia. As shown in FIGS. 5A-5D, between any two of the plurality of measurement periods T1 is a period T2 in which no measurement is performed. In some preferred embodiments, T2 is constant.

The replenishment mode can be started and stopped by applying the replenishment potential difference V2 and removing the replenishment potential difference V2, respectively, and the corresponding currents are represented by Ib. V2 is in the range of 0.1V to 0.8V, preferably a constant value in the range of 0.2V to 0.5V. In the replenishment mode, during the replenishment period t2 (t2 is in the range of 0 to T2), the replenishment potential difference V2 is applied between the auxiliary electrode Aux and the counter electrode R / C, and the voltage of the counter electrode R / C is increased. It becomes higher than the voltage of the auxiliary electrode Aux. During the replenishment mode, as shown in FIG. 4B, the switches S1 and S4 are in the open circuit state, the switches S2 and S3 are in the closed circuit state, the working electrode W is in the open circuit state, and the auxiliary electrode Aux is grounded. The counter electrode R / C is + V2. As a result, an oxidation reaction of Ag occurs at the counter electrode R / C, and AgCl is replenished to the counter electrode R / C by the replenishment amount. In the constant voltage circuit, the replenishment potential difference V2 is a constant voltage and the measured output current is Ib. In the present invention, the amount or value of the volume of AgCl (unit: "Couron", represented by the symbol "C") is defined by calculating the area under the current curve. Therefore, the amount of AgCl consumed in the measurement mode is Ia * T1, and the amount of AgCl replenished in the replenishment mode is Ib * t2. In such a case, the replenishment amount of AgCl can be controlled by adjusting the period t2 in which the potential difference V2 is applied. In other words, the replenishment amount is equal to or not equal to the consumable amount (almost as well as greater or more), assuming that AgCl on the counter electrode R / C is kept within a safe storage range. Including small).

In FIGS. 5A-5D, the horizontal axis represents time, the curve of V1 represents the application and removal of the measured potential difference V1, and the curve of V2 represents the application and removal of the supplementary potential difference V2. See FIG. 5A. In a preferred embodiment, both V2 and T2 are constant values, and the period t2 (ie, replenishment period) to which V2 is applied is a variable value. The replenishment period t2 is dynamically adjusted in the range of 0 to T2 according to the physiological signal Ia measured during the measurement mode and the measurement period T1. As shown in FIG. 5A, t2 is either t2 ′, t2 ″, or t2 ″ ″…. That is, the replenishment period t2 can be changed according to the amount of AgCl consumed. When the amount of AgCl consumed is large, the counter electrode R / C can be replenished for a long time in order to keep the AgCl of the counter electrode R / C within a safe storage range. For example, the amount of AgCl replenished during t2 ″ is greater than the amount of AgCl replenished during t2 ″.

With reference to FIG. 5B, in another preferred embodiment, V2, T2 and t2 are all constant values, where t2 = T2. That is, the measurement mode and the replenishment mode are seamlessly switched, and the period during which the measurement is not performed is the replenishment period. With reference to FIGS. 5C and 5D, in some preferred embodiments, V2, T2 and t2 are constant values, where t2 is a constant value greater than 0 and less than T2, eg, t2 =. 1/2 T2, 2/5 T2, 3/5 T2, and the like. The difference between FIG. 5C and FIG. 5D is that in FIG. 5C, after each measurement mode, before the replenishment mode starts, a buffer time (buffer time = T2-t2) elapses, and in FIG. 5D, each measurement mode After that, the replenishment mode starts immediately without buffering time, and there is a period between the end of each replenishment mode and the start of the next measurement mode. In some preferred embodiments, t2 is smaller than T2 and t2 can be any period in T2.

With reference to FIGS. 5E and 5F, FIGS. 5E and 5F show schematic current-time diagrams of constant voltage circuits operating alternately in measurement and replenishment modes in different ways. In FIGS. 5E and 5F, the horizontal axis represents time, the vertical axis represents current, and the curve represents a physiological parameter curve calculated from the measured physiological signal Ia. In the two embodiments, as in FIG. 5A, V2 and T2 are constant values and the replenishment period t2 is a variable value. In FIGS. 5E and 5F, the white area below the curve represents the AgCl consumption amount (Ia * T1) in the measurement mode, and the diagonal area represents the AgCl replenishment amount (Ib * t2) in the replenishment mode. Seen from this figure, the replenishment period t2 is from 0 to T2 according to the measured physiological signal Ia and the measurement period T1 in order to bring Ib * t2 closer to Ia * T1 or within a specific range of Ia * T1. It can be seen that it is dynamically adjusted within the range of. If necessary, the replenishment mode can be executed by selecting the front part (shown in FIG. 5E) or the rear part (shown in FIG. 5F) of the period (T2) in which the measurement mode is not executed.

Switching application of segment constant current circuit With reference to FIGS. 6A-6B and 8A-8C, FIGS. 6A and 6B show the segment constant current circuit in the measurement mode and the replenishment mode according to the present invention, respectively. 8A-8C show schematic voltage diagrams of constant current circuits operating in measurement mode and replenishment mode alternately in different ways, respectively. The measurement mode can be started and stopped by applying the measurement potential difference V1 and removing the measurement potential difference V1, respectively, and the corresponding current is represented by Ia. In the measurement mode, the measurement potential difference V1 is applied between the working electrode W and the counter electrode R / C during the measurement period T1. In the measurement mode, as shown in FIG. 6A, the switches S1 and S4 are in the closed circuit state, the remaining switches are in the open circuit state, the working electrode W is + V1, and the auxiliary electrode Aux is in the open circuit state. The counter electrode R / C is grounded. As a result, an oxidation reaction occurs at the working electrode W, and the working electrode W electrochemically reacts with the chemical reagent and the analyte to output the physiological signal Ia. AgCl in the counter electrode R / C has a consumption amount corresponding to the physiological signal Ia. As shown in FIGS. 8A-8C, between any two of the plurality of measurement periods T1 is a period T2 in which no measurement is performed. In some preferred embodiments, T2 is constant.

The replenishment mode can be started and stopped by applying a replenishment potential difference V2, which is a variable value, and removing the replenishment potential difference V2, and the corresponding current is represented by Ib. In the replenishment mode, the replenishment potential difference V2 is applied between the auxiliary electrode Aux and the counter electrode R / C during the replenishment period t2 (t2 is in the range of 0 to T2). During the replenishment mode, as shown in FIG. 6B, the switches S1 and S4 are in the open circuit state, and at least one of the switches corresponding to the switches S2 and I_F1 to I_Fn is in the closed circuit state (in FIG. 6B, I_F1 and I_Fn). The switch corresponding to I_F3 is in the closed circuit state), the working electrode W is in the open circuit state, the auxiliary electrode Aux is grounded, and the counter electrode R / C is + V2, and as a result, on the counter electrode R / C. , Ag oxidation reaction occurs, and AgCl is replenished to the counter electrode R / C. In the replenishment mode, depending on the magnitude of the physiological signal Ia and the measurement period T1, at least one of the switches corresponding to I_F1 to I_Fn can be turned on to output a constant current Ib, and AgCl can be replenished. The amount can be controlled by adjusting the period t2 in which the potential difference V2 is applied. That is, assuming that AgCl on the counter electrode R / C is kept within a safe storage range, the replenishment amount is equal to or not equal to the consumable amount (almost the same, larger or smaller). Including).

Switching application of continuously variable constant current circuit With reference to FIGS. 7A to 7B and 7A to 7B, FIGS. 7A and 7B are continuously variable constant current circuits according to the present invention in the measurement mode and the replenishment mode, respectively. Is shown. Since the measurement mode and the replenishment mode in this embodiment are the same as those in FIGS. 6A to 6B, they will not be described repeatedly here. This embodiment is different only in the replenishment mode of the embodiments of FIGS. 6A and 6B, and the constant current Ib can be output under the control of a digital-to-analog converter (DAC) based on the physiological signal Ia. The amount of AgCl replenished can be controlled by adjusting the period t2 in which the potential difference V2 is applied. That is, assuming that AgCl on the counter electrode R / C is kept within a safe storage range, the replenishment amount is equal to or not equal to the consumable amount (almost the same, larger or smaller). Including).

In FIGS. 8A-8C, the horizontal axis represents time, the vertical axis represents current, the curve of V1 represents the application and removal of the measured potential difference V1, and the curve of V2 represents the application and removal of the supplementary potential difference V2. As shown in FIG. 8A, in a preferred embodiment, T2 is a constant value and the period t2 (ie, the replenishment period) to which V2 is applied is a variable value. The replenishment period t2 is dynamically adjusted in the range of 0 to T2 according to the measurement period T1 and the physiological signal Ia measured in the measurement mode. As shown in FIG. 8A, t2 is either t2 ″, t2 ″, or t2 ″ ″. That is, the replenishment period t2 can be changed according to the amount of AgCl consumed. When the amount of AgCl consumed is large, the counter electrode R / C can be replenished for a long time in order to keep the AgCl of the counter electrode R / C within a safe storage range.

With reference to FIG. 8B, in another preferred embodiment, V2 is a variable value, T2 and t2 are constant values, where t2 is a constant value greater than 0 and less than T2. For example, t2 can be 1/2 T2, 2/5 T2, 3/5 T2, and so on. In this embodiment, V2 is dynamically adjusted according to the amount of AgCl consumed (ie, measurement mode) in the step of measuring the physiological signal. An example of the dynamic adjustment method is shown below. For example, a segment constant current circuit is used. This circuit includes n constant current power supplies and n switches, and each constant current power supply corresponds to a switch. In the replenishment mode, at least one of the n switches is turned on (closed circuit state) according to the amount of AgCl consumed, and a constant current value is output. When the replenishment period t2 is a constant value, the replenishment amount of AgCl can be controlled by selecting a different constant current output.

With reference to FIG. 8C, in another preferred embodiment, V2 is a variable value, T2 and t2 are constant values, where t2 = T2. That is, the measurement mode and the replenishment mode are seamlessly switched, and the period during which the measurement is not executed is the replenishment period.

Compared to a continuously variable constant current circuit, a segment constant current circuit can control multiple current paths with multiple switches, so it can be replenished in multiple segments according to the amount of current required. It can save power and reduce costs. Further, whether it is a constant voltage circuit or a constant current circuit, the potential difference can be generated from a DC power source or an AC power source, preferably a DC power source.

The embodiments of FIGS. 5A-8C all relate to an operation method in which a measurement step and a replenishment step are alternately repeated. This means that there is an AgCl replenishment step between any two measurement steps. Such a method makes it more certain that AgCl is kept within a safe storage range. However, in some preferred embodiments, Y rounds of AgCl replenishment can be optionally performed during the N measurement so that the cumulative amount of AgCl replenishment can still be kept within a safe storage range, where. Y ≦ N. The measurement and replenishment steps do not necessarily have to be performed in alternating cycles. The replenishment step can also be performed after several measurement steps or after a predetermined measurement time. For example, the replenishment step can be performed after 10 measurement steps or after the cumulative measurement time reaches 1 hour.

With reference to FIG. 8D, FIG. 8D shows a schematic current-time diagram of a constant current circuit operating alternately in measurement mode and replenishment mode in a manner similar to FIG. 8C. In FIG. 8D, the curve represents a physiological parameter curve calculated from the measured physiological signal Ia, and the conditions in which T2 and t2 are both constant values and V2 is a variable value are the same as those in FIG. 8C. is there. In FIG. 8D, the white area below the curve represents the amount of AgCl consumed in the measurement mode (Ia * T1), and the diagonal area represents the amount of AgCl replenished in the replenishment mode (Ib * t2). From this figure, the replenishment potential difference V2 may be dynamically adjusted according to the amount of AgCl consumed in order to bring Ib * t2 closer to Ia * T1 or within a specific range of Ia * T1. Understand.

Further, FIGS. 5E, 5F and 8D do not show the output timing of each physiological parameter value after executing each measurement step for measuring the physiological signal, but the physiological parameter value can be output, and the present invention is not limited thereto. The AgCl replenishment step can be performed, but not limited to, when the measurement is complete, or during the replenishment period, after all physiological parameters have been output, or after the physiological signal has been obtained.

In a two-electrode system that includes a working electrode W and a counter electrode R / C, the working electrode W needs to switch continuously between the execution of the oxidation reaction and the execution of the reduction reaction. In the chemical reaction environment of the electrode, switching between the oxidation reaction and the reduction reaction requires a stabilization period such as several seconds or several minutes. In contrast, in a three-electrode system containing a working electrode W, a counter electrode R / C and an auxiliary electrode Aux, a loop containing the working electrode W and the counter electrode R / C can be used in the measurement step with the auxiliary electrode Aux. The loop containing the counter electrode R / C can be used for the replenishment step. Therefore, the drawback that the working electrode W requires a stabilization period is avoided. That is, the replenishment step can be performed immediately after the measurement step.

With reference to FIG. 9, FIG. 9 shows a method of measuring an analyte according to the present invention. The service life of the microbiosensor can be extended by this method. The microbiosensor is, for example, the microbiosensor shown in FIGS. 2A to 3 and is to be implanted subcutaneously to measure a physiological signal representing a physiological parameter related to an analyte in a biological fluid (such as tissue fluid). Used for. In the embodiment of FIG. 9, the analyte may be glucose in tissue fluid, the physiological parameter is the glucose level of the human body, and the physiological signal is the current value measured by the microbial sensor. In this embodiment, the method for measuring the analyte comprises repeatedly performing the measurement step (S901) and the replenishment step (S902). In the measurement step (S901), the above-mentioned constant voltage circuit or the above-mentioned constant current circuit is used to execute the above-mentioned measurement mode during the measurement period T1 to output a physiological signal (that is, a current value), and at the same time, the counter electrode. AgCl in the above includes having a consumable amount corresponding to the current value. The measurement step (S901) also includes stopping the measurement step by stopping the measurement mode, and the current value is calculated to output a physiological parameter (ie, glucose level).

The chemical reaction formula in the measurement step (S901) is as follows.
The following oxidation reaction occurs at the working electrode 320.
The following reduction reaction occurs at the counter electrode 330.

The replenishment step (S902) includes performing the replenishment mode described above during the replenishment period using the constant voltage circuit or constant current circuit described above, so that AgCl on the counter electrode corresponds to the amount consumed. AgCl has a replenishment amount to allow, and therefore the amount of counter electrode is controlled within a safe storage range. As a result, the potential difference between the working electrode and the counter electrode can be kept stable, so that the obtained current value can maintain a stable correlation with the glucose value (when the detected substance is another analyte). , The correlation can be proportional or inversely correlated). In other words, it is possible to maintain a stable correlation between the next current value and the next glucose value obtained in the next measurement step. The replenishment step (S902) also includes a step of stopping the replenishment step by stopping the replenishment mode described above. After the replenishment step (S902) is completed, the method returns to the measurement step (S901) until the N measurement steps (S901) and the N replenishment steps (S902) are executed.

The chemical reaction formula in the replenishment step (S902) is as follows. The following reduction reaction occurs at the auxiliary electrode.
The positive potential of the counter electrode 330 causes the following oxidation reaction that occurs in the counter electrode 330.
Ag on the counter electrode is oxidized to Ag ^+, the body of a Cl ^- or from the oxidation of AgCl (or dissociation) Cl ^- bonded to form a AgCl with and, of AgCl was depleted during the measurement period T1 one Part or all is replenished to the counter electrode.

Humans can ingest chloride and iodide ions through iodine-doped salts. Available halogen ions include at least chloride and iodide ions to replenish the counter electrode with silver halide.

The following embodiments relate to a cycle of N measurement steps (S901) and N replenishment steps (S902). The physiological parameter referred to is preferably a glucose value and the physiological signal referred to is preferably a current value. According to some preferred embodiments, each measurement potential difference V1 is applied during the measurement period T1. Each replenishment potential difference V2 is applied during the replenishment period t2. The measurement period T1 is a constant value and can be a value within 3 seconds, 5 seconds, 10 seconds, 15 seconds, 30 seconds, 1 minute, 2.5 minutes, 5 minutes or 10 minutes. According to some preferred embodiments, the constant value may be a value within 30 seconds. The measurement period T1 is a constant value, and is 2.5 seconds, 5 seconds, 15 seconds, 30 seconds, 1 minute, 2.5 minutes, 5 minutes, 10 minutes, or 30 minutes, preferably 30 seconds. According to some preferred embodiments, each measurement period T1 plus each replenishment period t2 is a constant value. According to some preferred embodiments, each replenishment potential difference V2 has a constant voltage value and each replenishment period t2 is dynamically adjusted according to each consumption of AgCl (as shown in FIG. 5A). According to some preferred embodiments, the physiological parameters of each output are obtained through the calculation of physiological signals at a single measurement time point in each measurement period T1. According to some preferred embodiments, the physiological parameters of each output are obtained through mathematically calculated values of the plurality of physiological signals at the plurality of measurement points in each measurement period T1. The above-mentioned mathematically calculated values are, for example, a cumulative value, an average value, a median value, an average value of medians, and the like. According to some preferred embodiments, the replenishment amount of AgCl on the counter electrode is controlled so that each replenishment amount is equal to or not equal to each consumption amount (including almost the same, larger or smaller). Is controlled within a safe storage range. As a result, the next physiological signal obtained during the next measurement step maintains a stable proportional correlation with the next physiological parameter. According to some preferred embodiments, the step of removing each measured potential difference V1 is to disconnect the circuit connecting the working electrode and the counter electrode or set each measured potential difference V1 to zero. That is, the power can be turned off to open the measurement circuit, or a zero volt voltage can be applied between the working electrode and the counter electrode, where the operating time of either of the two operations Is 0.01 to 0.5 seconds. By removing the measurement potential difference V1, the generation of the Λ-type physiological signal can be avoided. According to some preferred embodiments, the step of removing each replenishment potential difference V2 is to either disconnect the circuit configured to connect the auxiliary electrode to the counter electrode or set each replenishment potential difference V2 to zero. Is.

According to some preferred embodiments, after the biosensor is implanted in the human body, the biosensor is in equilibrium and stability in the body to stably present a physiological signal that is positively correlated with the concentration of the analyte. , Need warm-up time. Therefore, in the measurement step (S901), the measurement voltage is continuously applied until the end of the measurement period T1, and the measurement period T1 is controlled so that the physiological signal and the physiological parameter of the analyte have a stable proportional correlation. .. For this purpose, the measurement period T1 can be a variable value, or a combination of variable and constant values (eg, variable + constant, where the variable values are 1 hour, 2 hours, 3 hours, 6). Hours, 12 hours or 24 hours, with constant values being, for example, 30 seconds).

With reference to FIGS. 5A-5F, 8A-8D and 9, the present invention measures the combined current of the counter electrode using the voltage applied to the counter electrode R / C during a period of time to determine the initial AgCl. The capacitance is obtained by mathematically calculating the combined current during the period. For example, the initial capacitance of AgCl is defined by calculating the area under the curve of the combined current. _{The initial volume of AgCl is also referred to as the initial} amount or the initial amount of Coulomb (Cinitial), and the following are all expressed as amounts. The counter electrode R / C contains Ag and AgCl. If the amount of AgCl (X% AgCl) is known, the amount of Ag can be calculated (Y% Ag = 100% to X% AgCl). In each measurement step (S901), the amount of AgCl consumed ( _{indicated by C consume} ) is defined by calculating the area of the working electrode W under the current curve. AgCl of the counter electrode R / C has a consumed amount (C _consume ) corresponding to the physiological signal Ia, that is, C _consume = Ia * T1. In each replenishment step (S902), each replenishment amount of AgCl ( _{indicated by Creplenish} ) is defined by calculating the area of the counter electrode R / C under the current curve. _{That, C replenish} = Ib * _t2, where, t2 is in the range of 0～T2.

The calculation method of the safe storage tube amount of AgCl is shown below. In some preferred embodiments, the safe storage range is represented by the Ag to AgCl ratio. The present invention uses the amount of Coulomb (C) measured at the counter electrode to reflect the Ag to AgCl ratio. In some preferred embodiments, the Ag to AgCl ratios are 99.9%: 0.1%, 99%: 1%, 95%: 5%, 90%: 10%, 70%: 30%, 50. %: 50%, 40%: 60% or 30: 70%, which guarantees that a certain amount of AgCl is not exhausted on the counter electrode, so that each measurement step for measuring the physiological signal. Can be executed stably. The residual amount of AgCl is the sum of the replenishment amount and the initial amount minus the consumption amount. In some preferred embodiments, the residual amount of AgCl varies within a range, i.e., the residual amount of AgCl is controlled within a range in which a specific value (X value) is increased or decreased from the initial amount. That is, ( _Creplenish + _Cinitial ) -C _context = _Cinitial ± X, where 0 <X <100% _Cinitial , 10% _Cinitial < _X≤90 % Cinitial, or 0.5% _Cinitial <X≤ It is 50% _Clinical . In some preferred embodiments, the residual amount of AgCl can be in the range, gradually decreasing, gradually increasing, steadily changing, or optionally changing, but still within the range.

With reference to FIG. 10, FIG. 10 shows a method for measuring an analyte according to another embodiment of the present invention. By this method, the service life of the microbiosensor can be extended, and the amount of silver and silver halide materials of the counter electrode can be reduced. The microbiosensor is, for example, the microbiosensor shown in FIGS. 2A to 3 for being implanted subcutaneously to measure a physiological signal representing a physiological parameter associated with an analyte in a biological fluid (such as tissue fluid). used. The electrode material of the counter electrode of the microbiosensor contains silver and silver halide. In the embodiment of FIG. 10, the analyte is glucose in tissue fluid, the physiological parameter is the glucose level of the human body, and the physiological signal is the current value measured by a microbiosensor. Only one cycle of this embodiment will be described below. The method of this embodiment begins with the step of applying a measuring voltage to drive a working electrode and measuring a physiological signal to obtain physiological parameters, where a specific amount of silver halide is consumed (hereinafter,). "Consumable amount") (S1001).

Next, the step of applying the measurement voltage is stopped (S1002), and the obtained physiological signal is used to acquire the physiological parameters (S1003). After acquiring the physiological parameters, a replenishment voltage is applied between the counter electrode and the auxiliary electrode to drive the counter electrode so that silver halide is replenished by the replenishment amount (S1004). Here, the value obtained by subtracting the consumable amount from the sum of the replenishment amount and the initial amount (that is, the above-mentioned "residual amount") is controlled within a range in which a specific value is increased or decreased from the initial amount. The above control step is by controlling the replenishment amount to be equal to or not equal to the consumable amount (including approximately the same, more, or less) in order to keep the amount of silver halide within a safe storage range. Achieve. According to the chemical reaction formula, an increase or decrease in the number of moles of silver halide corresponds to an increase or decrease in the number of moles of silver. Therefore, for simplicity of explanation, the amount of silver halide consumed corresponds to the simulated increase in silver. In some preferred embodiments, the value of the remaining amount is controlled so that the ratio of the amount of silver halide to the sum of the amount of silver halide and the amount of silver (AgCl / Ag + AgCl) is greater than zero and less than one. (That is, the counter electrode must contain a certain amount of silver halide). The ratio is preferably between 0.01 and 0.99, 0.1 to 0.9, 0.2 to 0.8, 0.3 to 0.7 or 0.4 to 0.6. .. When the replenishment amount is reached, the step of applying the replenishment voltage is stopped (S1005). Next, the method returns to step S1001 and executes the next loop.

Specific embodiments of the present invention will be described below. As an example, the useful life of a biosensor needs to reach 16 days. For this purpose, a method of calculating the required size of Ag / AgCl material on the signal sensing portion of the electrode will be described below. For example, the average measurement current of the analysis target of each measurement is 30 nA, the measurement period (T1) is 30 seconds, and the replenishment period (t2) is 30 seconds. _{The daily consumption} of AgCl (Cconse / day) = 1.3 mC / day. Assuming that the biosensor service life requirement is 16 days, the amount of AgCl consumed to use 16 days is 1.3x16 = 20.8 mC.

For example, the length of the counter electrode is 2.5 mm, which _{corresponds to the initial} amount of AgCl, Clinical = 10 mC.
(1) When the service life of the sensor is 16 days without replenishing AgCl, the required length of the counter electrode is at least as follows.
C _{16 day / C context} / day = 20.8 mC / 1.3 mg / day = 16 mm.
(2) Therefore, if the silver halide replenishment method in the present application is not carried out, the length of the counter electrode needs to exceed 16 mm in order to make the service life of the sensor 16 days.

In this embodiment, provided that the silver halide replenishment technique of the present invention is not used, the counter electrode signal detector is made of a relatively large size Ag / AgCl material in order to achieve a service life of 16 days. Need to be configured. According to the silver halide replenishment method in the present invention, the silver halide replenishment step is performed between the two measurement steps. Since the silver halide cycle is consumed and replenished in a short time (replenished at the time of use), the amount of Ag / AgCl material in the sensor can be reduced, which makes the sensor smaller. Therefore, it is necessary to prepare a 16-day AgCl capacity for the material of the signal sensing portion of the electrode and consume it. For example, if the AgCl volume is prepared for about 1-2 days, the sensor usage time will be 16 days. Therefore, the present invention has the effect of extending the service life of the sensor. The volume of AgCl for 1-2 days also refers to the initial amount of AgCl in the counter electrode before factory shipment or before performing the first measurement. The initial amount of AgCl can be, for example, between about 1.3 and 2.6 mC, but can be in other smaller or larger ranges. In other embodiments, different AgCl volumes of 1-5 days, 1-3 days, 6-24 hours, and 6-12 hours can also be prepared. The size of the signal detection portion of the counter electrode should be configured so that the counter electrode has a capacity that allows each measurement step of glucose to be performed stably and a positive correlation between the measured current and the glucose concentration in the body. Can be done.

The prior art increases the length / area of the electrodes so that the sensor reaches the required measurement date without using the silver chloride replenishment technique of the present invention. For example, the length of the embedded end of the prior art is about 12 mm. Due to the long implant length of the prior art, the implant must be implanted subcutaneously at an oblique angle to prevent the implant from being deeply implanted in the subcutaneous tissue and causing a large implant wound. As another example, the volume of AgCl for 1-2 days is converted to about 1.3-2.6 mC, and the length of the counter electrode for 1-2 days is converted to 2.5-5 mm. Therefore, if the silver halide replenishment method of the present invention is not used, the length of the counter electrode is required to be 16 mm. Compared to the above example, it is clear that the present invention has a more important effect on reducing the size of the counter electrode. According to the silver chloride replenishment step of the present invention, the embedded end of the present invention can be shortened to, for example, 10 mm or less. With reference to FIGS. 2A-2C, the lower half of the connection region 317 to the second end 314 of the microbiosensor 300 of the present invention forms a short embedded end 318 as shown in FIGS. 2A-2B. The embedding depth of the short embedding end 318 is at least the depth to the dermis where the glucose concentration in the tissue fluid can be measured. According to the silver chloride replenishment step of the present invention, the length of the longest side of the short embedding end 318 is 6 mm or less, so that the short embedding end 318 of the microbiosensor 300 is vertically embedded under the biological epidermis. be able to. Preferably, the length of the longest side of the short embedded end 318 is 5 mm, 4.5 mm, 3.5 mm, or 2.5 mm or less. The short embedded end 318 of the present invention includes the signal sensing portion 332 of the counter electrode 330, and the length of the longest side of the signal sensing portion 332 of the counter electrode 330 is 6 mm or less, preferably 2 to 6 mm, 2 to 5 mm, 2 It is ~ 4.5 mm, 2 to 3.5 mm, 0.5 to 2 mm, or 0.2 to 1 mm.

Therefore, as compared with the case where the silver halide replenishment technique of the present invention is not used, the silver halide replenishment method of the present invention can effectively extend the service life of the microsensor and Ag / AgCl on the counter electrode. The use of material can be significantly reduced and the size of the counter electrode signal detector is reduced. By reducing the use of Ag / AgCl material in the counter electrode, the sensor can be miniaturized and biotoxicity can be reduced. In addition, reducing the size of the electrodes specifically refers to the shortened length of the embedded end of the sensor, which will reduce the user's pain during implantation.

Although the present invention has been described above using the embodiments, the technical scope of the present invention is not limited to the scope described in the above embodiments. It will be apparent to those skilled in the art that various changes or improvements can be made to the above embodiments. It is clear from the description of the claims that such modified or improved forms may also be included in the technical scope of the present invention.

Physiological signal measuring device 10
User device 20
Transmitter 200
Processor 210
Power supply 220
Circuit switching unit 230
Temperature sensing unit 240
Communication unit 250
Microbiosensor 300
Board 310
Surface 311
Surface 312
First end part 313
Second end 314
Signal output area 315
Sensing area 316
Contiguous zone 317
Embedded end 318
Working electrode 320, W
Signal output part 321
Signal sensing part 322, 332, 342
First working electrode 323
Second working electrode 324
Third working electrode 325
Counter electrode 330, R / C
Auxiliary electrode 340, Aux
Chemical reagent 350
Physiological signals Ia, Ib
Switches S1, S2, S3, S4, I_F1 to I_Fn
Measurement period T1
Measurement potential difference V1
Replenishment potential difference V2
Replenishment period t2

Claims (20)

A biosensor implanted subcutaneously to measure a physiological signal representing a physiological parameter associated with an analyte in a biofluid, comprising a working electrode, a counter electrode and an auxiliary electrode, wherein the working electrode is the same as the analyte. The counterelectrode is at least partially covered with a chemical reagent to react and the counterelectrode is measured using the biosensor to extend the life of the biosensor with silver and silver halide. It ’s a method,
(A) A measurement step in which a measurement is performed.
i, In order to cause a first oxidation reaction at the working electrode having an electrochemical reaction with the chemical reagent and the analysis substance and output the current physiological signal, the working electrode of the counter electrode during the measurement period In the step of applying a measured potential difference between the working electrode and the counter electrode so as to have a voltage level higher than the voltage level, the silver halide of the counter electrode corresponds to the current physiological signal. Substeps with current consumption and
ii, the measurement step including the sub-step of removing the measurement potential difference, stopping the measurement step, calculating the current physiological signal and outputting the current physiological parameter.
(B) A replenishment step for executing replenishment.
i, the replenishment potential difference between the counter electrode and the auxiliary electrode is applied during the replenishment period so that the counter electrode has a voltage level higher than the voltage level of the auxiliary electrode. The silver dioxide causes a dioxide reaction in the silver, so that the halide silver obtains a replenishment amount corresponding to the consumable amount, the halogenated silver of the counter electrode has an amount maintained in a safe storage range, and then The sub-step that keeps the next physiological signal and the next physiological parameter obtained in the measurement step in a specific correlation, and
ii, the replenishment step including a sub-step that removes the replenishment potential difference and stops the replenishment step.
(C) A step of executing the next measurement step including the same substep as step (a), and
(D) A method comprising: (d) a step of executing the next replenishment step including the same sub-step as step (b). The measured potential difference and the replenishment potential difference are applied to the measurement period and the replenishment period, respectively.
The method according to claim 1, wherein the measurement period has a time value that is either a constant measurement period value or a variable measurement period value. The method according to claim 2, wherein the total period of the measurement period and the replenishment period is a constant. The replenishment potential difference has a constant voltage value and
The method according to claim 2, wherein the replenishment period is dynamically adjusted based on the consumption amount of the silver halide. The replenishment period has a constant period value and
The method according to claim 2, wherein the replenishment potential difference has a value that is dynamically adjusted based on the consumption amount of the silver halide. The constant measurement period value is not more than or equal to the time value selected from the group consisting of 3 seconds, 5 seconds, 10 seconds, 15 seconds, 30 seconds, 1 minute, 2 minutes, 5 minutes, and 10 minutes. The method according to claim 2. The first aspect of the present invention is characterized in that the amount of the silver halide of the counter electrode within the safe storage range is maintained by controlling the replenishment amount to be close to or equal to the consumption amount. the method of. The first aspect of the present invention is characterized in that the amount of the silver halide of the counter electrode within the safe storage range is maintained by controlling the replenishment amount to be larger than the consumption amount. the method of. The first aspect of the present invention, wherein the amount of the silver halide of the counter electrode within the safe storage range is maintained by controlling the replenishment amount to be less than the consumption amount. Method. The first aspect of the present invention, wherein the amount of the silver halide of the counter electrode within the safe storage range is maintained by controlling the replenishment amount so as not to be equal to the consumption amount. Method. A biosensor implanted subcutaneously to measure a physiological signal representing a physiological parameter associated with an analyte in a biofluid, comprising an working electrode, a counter electrode and an auxiliary electrode, wherein the working electrode is at least by a chemical reagent. Partially covered, the counterelectrode is a method of measuring the analyte using the biosensor to extend the useful life of the biosensor, which contains silver and silver halide and has an initial amount. ,
A step of acquiring the physiological parameter by applying a measuring voltage to drive the working electrode and measuring the physiological signal, and a step of consuming the silver halide by the amount of consumption.
The step of stopping the application of the measurement voltage and
Each time the physiological parameter is acquired, a replenishment voltage is applied between the counter electrode and the auxiliary electrode to drive the counter electrode and cause an oxidation reaction to bring a replenished amount of the silver halide to the counter electrode. With steps to replenish,
A method characterized in that a guard value obtained by subtracting the consumable amount from the total of the replenishment amount and the initial amount is controlled within a range in which a specific value is increased or decreased from the initial amount. The measured voltage is applied during the measurement period and
The replenishment voltage is applied during the replenishment period and
11. The method of claim 11, wherein the measurement period has a time value that is either a constant measurement period value or a variable measurement period value. The supplementary voltage has a constant voltage value and has a constant voltage value.
12. The method of claim 12, wherein the replenishment period is dynamically adjusted based on the consumable amount of the silver halide. The replenishment period has a constant period value and
12. The method of claim 12, wherein the replenishment voltage has a value that is dynamically adjusted based on the amount of consumption of the silver halide. In order to keep the amount of silver halide in a safe storage range, the replenishment amount of the silver halide is selected from the group close to, equal, large, small, unequal to the consumable amount of the silver halide. The method according to claim 11, wherein a guard value is secured by controlling the amount of silver halide. The specific value is X
The method according to claim 11, wherein X satisfies the condition of 0 <X <100% of the initial amount. A microbiosensor for subcutaneous implantation for measuring physiological parameters that represent physiological signals associated with an in vivo analyte.
With the board
With chemical reagents
A working electrode placed on the substrate, at least partially covered with the chemical reagent, driven for a first oxidation reaction within the measurement period, to measure the physiological signal and obtain the physiological parameters.
A pair electrode arranged on the substrate and containing silver and silver halide, wherein the silver halide has an initial amount and is consumed at a specific consumption amount within the measurement period.
Auxiliary electrodes arranged on a substrate, each time the physiological parameters are acquired, the counter electrode and the auxiliary electrode are driven for the carbon dioxide reaction within the replenishment period, and the replenishment amount of the halogen The auxiliary electrode, in which silver chemicals are replenished to the counter electrode, is provided.
A microbiosensor characterized in that a guard value obtained by subtracting the consumable amount from the total of the replenishment amount and the initial amount is controlled within a range in which a specific value is increased or decreased from the initial amount. The guard value according to claim 17, wherein the ratio of the amount of silver halide to the sum of the amount of silver and the amount of silver halide is controlled to be larger than zero and smaller than 1. Microbiosensor. The microbiosensor according to claim 17, further comprising a short embedded end having a length of 6 mm or less. The microbiosensor according to claim 17, wherein the counter electrode is at least partially covered with the chemical reagent, and the auxiliary electrode contains platinum.